The aim is to identify and study the mechanisms by which volatile anesthetics depress contractility of heart muscle in vitro. The approach will use technology that allows one to evaluate simultaneously variables of mechanical function (length, force, velocity, elastic stiffness and time) and changes in intracellular calcium concentration detected with the Ca2+-sensitive bioluminescent protein aequorin in intact living ventricular cardiac muscle. In the first project the hypothesis will be tested that volatile anesthetics halothane, enflurane and isoflurane do not alter (1) the luminescent reaction of aequorin with Ca2+ ions, i.e., quantum yield and Ca2+ sensitivity of the Ca2+-regulated bioluminescent protein aequorin, and the apparent association constants of various metal chelators (EGTA, EDTA, CDTA) for Ca2+. This information will serve to quantify light signals from aequorin into intracellular calcium concentrations in living heart muscle. Three detailed studies of the intracellular free calcium concentration transient in rhythmically contracting heart muscle will allow one to pinpoint effects of each of the volatile agents on (1) release and entry of Ca2+ in the cytoplasm, (2) uptake of Ca2+ into intracellular storage sites, (3) apparent sensitivity of the contractile proteins for Ca2+, and (4) influx of Ca2+ through the cardiac sarcolemmal membrane. This approach has the advantage that it assesses the relative contribution of several cellular components involved in excitation-contraction coupling in an unperturbed, intact muscle. In a fifth study, instantaneous elastic stiffness will be measured throughout contraction and relaxation of heart muscle in the absence and presence of anesthetic agents. The information will shed light on the time course of onset and decay of instantaneous force potential of heart muscle in the presence of volatile anesthetics. The relative dissociation in time of the decline of the calcium transient and of elastic stiffness will provide corroborative information of the effects of anesthetics on the detachment of crossbridges.